The purpose of this project is to study cholesterol efflux from human monocyte-derived macrophages, a major cell type that accumulates cholesterol in atherosclerotic lesions. We have found that incubation of monocyte-derived macrophages with nonlipoprotein cholesterol leads to massive accumulation of unesterified and esterified cholesterol. When cholesterol-loaded, monocyte-derived macrophages were incubated in serum-free medium, their total and unesterified cholesterol content decreased over a 72-hour period while their esterified cholesterol content remained the same or increased. This demonstrates that monocyte-derived macrophages have a means to secrete cholesterol even in the absence of a cholesterol acceptor such as HDL. Currently, we are characterizing the nature of the cholesterol secreted by monocyte-derived macrophages. Monocyte-derived macrophages secrete cholesterol (entirely unesterified) that is associated with two lipoprotein particles of densities 1.06 and 1.09 g/ml. Both lipoprotein particles demonstrate a cholesterol to phospholipid molar ratio of >1 and have different associated protein species. Studying cholesterol efflux in monocyte-macrophages is important to our understanding of why cholesterol accumulates within these cells in atherosclerotic lesions. Examination of the nature of the cholesterol-rich particles released from these cells and comparison of this to those cholesterol-rich particles we have isolated from human atherosclerotic lesions will help determine the origin of cholesterol that accumulates in lesions. These studies will also help to determine whether macrophages play a beneficial or adverse role in the pathogenesis of atherosclerosis.